Centrifugal microfluidics deals with handling of liquids within the pl to ml ranges in rotating systems. Such systems are mostly disposable polymer cartridges used in or instead of centrifuge rotors, with the intention of enabling completely novel processes which cannot be performed by manual processes or pipetting robots because of the precision or volume involved, or of automating laboratory processes. In this context, standard laboratory processes such as pipetting, centrifuging, mixing or aliquoting may be implemented in a microfluidic cartridge. To this end, the cartridges contain channels for directing fluids as well as chambers for collecting liquids. The cartridges are subject to a predefined sequence of rotary frequencies, the frequency protocol, so that the liquids contained within the cartridges may be directed into corresponding chambers by inertial forces. Centrifugal microfluidics is mainly applied in laboratory analytics and in mobile diagnostics.
The implementation of cartridges that has been most common up to now are centrifugal-microfluidic disks which are known, e.g., by the names or brands of “Lab-on-a-disk”, “Lab-Disk” and “Lab-on-CD”, which are inserted into specific processing equipment. Other formats such as a microfluidic centrifuge tube, which is known by the name of “LabTube”, may be inserted into rotors of already existing standard laboratory equipment.
One essential fundamental operation that is performed in centrifugal-microfluidic cartridges is retaining and releasing liquids in a calculated manner. The problem consists in transferring liquids from a first fluid chamber into a second fluid chamber at defined rotary frequencies or defined changes in the rotary frequencies, and/or in retaining liquids within a first chamber at defined rotary frequencies or defined changes in the rotary frequencies. For using said fundamental operation in a potential product, robustness of the process is paramount. Moreover, the fundamental operation should be implemented as a monolithically integrated valve so that no additional components or materials—which considerably increase the cost of the cartridge in terms of cost of materials or in terms of additional structural design and connection technology (assembly)—are required.
Monolithically integrated valves in centrifugal-microfluidic systems are known from conventional technology. For example, in “Pneumatic Pumping in Centrifugal Microfluidic Platform”, Microfluid Nanofluid, 2010, 9, pp. 541 to 549, R. Gorkin et al. describe a method of pneumatic pumping which enables retaining liquid within a first fluid chamber during a first phase at defined, high rotary frequencies (typically several 10 Hz) and to subsequently direct the liquid into a second fluid chamber during a second phase at defined, lower rotary frequencies. This involves transferring liquid from a reservoir into a first fluid chamber when the rotary frequency increases. When the rotary frequency increases, the liquid is retained within the first fluid chamber, a gas volume entrapped within the first fluid chamber being compressed. When the rotary frequency decreases, the entrapped gas volume expands again and displaces some of the liquid into a curved channel acting as a siphon. Once the siphon crest has been passed, an additional centrifugal pressure arises which causes the liquid to be transferred from the first into the second fluid chamber. Thus, a gas volume within the first fluid chamber that is entrapped by the process liquid is compressed during the first phase so as to make use, during the second phase, of the corresponding expansion of the gas volume for returning the liquid.
In the process of pneumatic pumping, a specific threshold value of the rotary frequency (threshold frequency) will be exceeded during the first phase so as to retain the liquid within the first fluid chamber. This very threshold frequency will subsequently be fallen below so as to return the liquid via the siphon crest and to start the transfer of fluid from the first fluid chamber into the second fluid chamber. In order for the filling of the siphon to be independent of capillary forces, the threshold frequency should be as high as possible.
S. Zehnle, F. Schwemmer, G. Roth, F. von Stetten, R. Zengerle and N. Paust, “Centrifugo-dynamic Inward Pumping of Liquids on a Centrifugal Microfluidic Platform”, Lab Chip, 2012, 12, pp. 5142 to 5145, describe a method of centrifugo-dynamic inward pumping which enables retaining liquid within a first fluid chamber at defined, high rotary frequencies (typically several 10 Hz) during a first phase and to subsequently direct a major part of the liquid into a second, radially inwardly located fluid chamber at a rapidly decreasing rotary frequency during a second phase. This involves transferring liquid from a reservoir into a first fluid chamber when the rotary frequency increases. When the rotary frequency increases, the liquid is retained within the first fluid chamber, a gas volume entrapped within the first fluid chamber being compressed. When the rotary frequency rapidly decreases, the entrapped gas volume expands again and displaces the major part of the liquid through that channel which has the smaller flow resistance. Thus, a gas volume within the first chamber that is entrapped by the process liquid is compressed during the first phase so as to make use, during the second phase, of the energy of the compressed gas for pumping the liquid radially inward. A corresponding method is described in DE 10 2012 202 775 A1.
As was set forth above, the threshold frequency should be as high as possible in pneumatic pumping so as to keep the influence of capillary forces low. This means that the siphon is typically also filled at high rotary frequencies (even if the delay rate amounts to several 10 Hz/s). The inventors have found that this has drawbacks. When the liquid reaches the siphon crest at relatively high rotary frequencies, this may cause instability of the liquid/gas interface at the siphon crest. Inclusion of air bubbles and, thus, function failure of the siphon may result. This effect might be minimized in a siphon having a small cross-sectional area, which would increase, however, the dependency on capillary forces as well as the fluidic resistance and, thus, the length of time during which the fluid transfer takes place. When liquid is pumped through a siphon at relatively high rotary frequencies, instability of the liquid/gas interface at the outer siphon end may also result. Here, too, inclusion of air bubbles and, thus, function failure of the siphon may be the consequence. Depending on the configuration of the siphon, the pressure in the siphon crest may become so low, in case of a high rotary frequency, that the liquid will evaporate and that consequently, formation of gas bubbles will result in a function failure of the siphon. Even at relatively low rotary frequencies and, thus, relatively low negative pressures, gas bubbles may form since, due to the relatively low pressure within the crest area of the siphon, the solubility of gases such as oxygen, for example, will decrease and consequently, the amount of gas which is no longer soluble will outgas in the form of bubbles.
If inward pumping as is described, e.g., in DE 10 2012 202 775 A1 is used as a function of a valve, this is disadvantageous in that it will never be the entire volume of liquid that will be transferred from the compression chamber into the collection chamber.
A further possibility of retaining liquids is to exploit capillary force which, controlled by the rotary frequency, is overcome by the centrifugal force in order to move the liquid. However, such methods are highly dependent on the surface tension of the liquid and on the nature of the surfaces of the fluidic channels and can therefore not be considered to be robust.